Introduction

SAFC® has developed a serum-free medium, EX-CELL 302, which meets or exceeds requirements for use with CHO-K1 cells grown in suspension culture. Raw materials (amino acids, lipid supplements and carbohydrates) used in this medium are of the highest quality: USP, EP, JP or ACS, and are non-animal sourced wherever possible. With the addition of 4 mM L-glutamine, EX-CELL 302 will support growth of CHO-K1 cells in batch suspension cultures for greater than eight (8) days while maintaining high cell viability (> 70%). This long culture period and stationary phase allow for an extended production period and higher culture productivity. L-glutamine, hypoxanthine and thymidine have been omitted from EX-CELL 302 allowing for use with either the DHFR- or GS selection systems.

Materials and Methods

CHO-K1 cells (ATCC CCL 61) were obtained from the American Type Culture Collection (Rockville, MD) and were previously adapted to EX-CELL 301 (Catalog No. 14331C - Available for custom development only).

Results

Figure 1. EX-CELL 302 CHO-K1 Growth Profile
(10 passages)CHO-K1 cultures were monitored daily for growth and viability. These data represent the average cell density and viability for 10 passages. Cultures were seeded at a density of 2 x 105 cells/mL in 100 mL media using a spinner speed of 60 rpm. An increase in growth rate and final cell density were seen in subsequent passages after full adaptation.

Conclusions

Data presented here demonstrate extended cellular growth in EX-CELL 302 medium for eight (8) days in spinner batch cultures. Cultures of CHO-K1 cells achieved an average maximum cell density of approximately 3.5 x 106 cells/mL with exponential doubling times in the range of 20 hours while maintaining viabilities greater than 70%. EX-CELL 302 contains low levels (< 100 µg/L total) of recombinant proteins. CHO-K1 cells, cryopreserved and adapted to serum-free growth in EX-CELL 301, were transferred directly into EX-CELL 302 without an adaptation period. SAFC's regulatory friendly EX-CELL 302 is suitable for the production of human therapeutic products since it does not contain animal-derived proteins or other undesirable components.